Inhibin is an ovarian-produced protein that regulates pituitary FSH release and follicle function. We hypothesize that the ability of specific cells in the reproductive tract to respond to inhibin is controlled by a specific inhibin receptor present on the cell membrane. Therefore, we will identify the inhibin receptor. The inhibin receptor has been partially purified from the gonadal tumors that arise from inhibin-deficient mice (alpha-subunit knockout mice). Using affinity purification techniques, inhibin receptor proteins have been purified. They have been labeled inhibin receptor based on the facts that they were membrane associated and bound inhibin A and not activin A. Moreover, the proteins are not follistatin. Based on this preliminary data, the current grant proposes to continue these studies and to purify and then characterize the inhibin receptor. Several alternative approaches to the identification of the receptor are also proposed. Receptor activation stimulates cytoplasmic signaling molecules. Candidate signal proteins have been identified (Smad2 and Smad4) and we have shown that these proteins are activated by inhibin in granulosa cells. We hypothesize that normal, inhibin-regulated granulosa cell function is dependent on the appropriate interaction of ligand with Smad2 or Smad4 and that alteration in the inhibin signal transduction pathway will lead to abnormal follicle function. These tenets will be tested in granulosa cell culture. The transcriptional response of inhibin-regulated promoters will be measured following over-expression of normal or mutated signal proteins in a granulosa cell line, GRM02. The genetic sequence that is responsive to inhibin signals will be identified. These studies will establish the causal relationship between the inhibin system and normal and abnormal follicle function and lead toward a more complete understanding of key regulating protein in the ovarian-pituitary axis.